Comparison of electron paramagnetic resonance methods to determine distances between spin labels on human carbonic anhydrase II.
Comparison of electron paramagnetic resonance methods to determine distances between spin labels on human carbonic anhydrase II.
复制标题
比较电子顺磁共振方法以确定人碳酸酐酶 II 上自旋标记之间的距离。
DOI：
10.1016/s0006-3495(01)76254-6
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发表时间：
2001
影响因子：
3.4
通讯作者：
Eaton,SS
中科院分区：
文献类型：
--
作者：
Persson,M;Harbridge,JR;Hammarström,P;Mitri,R;Mårtensson,LG;Carlsson,U;Eaton,GR;Eaton,SS
通过定点自旋标记法制备了四种双自旋标记的人碳酸酐酶II变体和相应的单自旋标记变体。自旋标签之间的距离，从连续波电子顺磁共振光谱通过分析的相对强度的半场过渡，傅立叶反卷积的线形加宽，和计算机模拟的线形变化。距离也由四脉冲双电子-电子共振测定。对于每种变体，至少有两种方法适用，并且方法之间具有合理的一致性。距离从7到24米不等。由于标记不完全，双自旋标记样品中含有一些单标记蛋白。比较了每种距离确定方法对非相互作用分量的灵敏度。
Four doubly spin-labeled variants of human carbonic anhydrase II and corresponding singly labeled variants were prepared by site-directed spin labeling. The distances between the spin labels were obtained from continuous-wave electron paramagnetic resonance spectra by analysis of the relative intensity of the half-field transition, Fourier deconvolution of line-shape broadening, and computer simulation of line-shape changes. Distances also were determined by four-pulse double electron-electron resonance. For each variant, at least two methods were applicable and reasonable agreement between methods was obtained. Distances ranged from 7 to 24Å. The doubly spin-labeled samples contained some singly labeled protein due to incomplete labeling. The sensitivity of each of the distance determination methods to the non-interacting component was compared.